(+-)-9-(beta-(3-alpha-amino-2alpha-hydroxy)cyclopentyl)-6-substituted-purines and derivatives thereof

ABSTRACT

A NEW CLASS OF CHEMICAL COMPOUNDS, ($)-9-(B(3AAMINO-2A-HYDROXY)CYCLOPENTYL) - 6 - SUBSTITUTED - PURINES HAVING THE GENERAL FORMULA:   6-R,9-(2-(HO-),3-X-CYCLOPENTYL)-9H-PURINE   WHEREIN R IS A SUBSTITUENT SELECTED FROM THE GROUP CONSISTING OF HYDROXYL, MERCAPTO, METHYL MERCAPTO, HALOGEN (CHLORINE, BROMINE, FLUORINE AND IODINE) AND SUBSTITUTED AMONO:   R&#39;&#39;-N(-R&#34;)-   WHEREIN R&#39;&#39; AND R&#39;&#39;&#39;&#39; MAY BE THE SAME OR DIFFERENT SUBSTITUENTS AND ARE SELECTED FROM THE GROUP CONSISTING OF HYDROGEN, METHYL, ETHYL, PROPYL AND PHENYL; X IS AN AMINO RADICALOR AMINO ACID; AND ACID SALT DERIVATIVES OF THE SAME, SUCH AS ACETIC, HYDROCHLORIC, SULFURIC, ETC.; AND THE METHOD OF PREPARATION THEREOF. THE INTERMEDIATE ($)-9(B-(3A-AMINO - 2A - HYDROXY)CYCLOPENTYL) - 6 - DIMETHYLAMINOPURINE, OR ANY OF ITS CORRESPONDING 6-SUBSTITUTED DERIVATIVES, LEADS TO THE PREPARATION OF TWO DIASTERIOISOMERS WHEN CONDENSED WITH THE APPROPRIATELY BLOCKED LAMINO ACID. THE SEPARATION OF THESE DIASTERIOISOMERS RESULTS IN THE ISOLATION OF ANTIMICROBIAL AND ANTITUMOR AGENTS.

United States Patent Otce 3,825,541 Patented July 23, 1974 U.S. Cl.260-253 4 Claims ABSTRACT F THE DISCLOSURE A new class of chemicalcompounds, *)-9-[,B-(3aamino-Za-hydroxy)cyclopentyl] 6 substitutedpurines having the general formula:

wherein R is a substituent selected from the group consisting ofhydroxyl, mercapto, methyl mercapto, halogen (chlorine, bromine,fluorine and iodine) and substituted amino:

Rl N \Rll wherein R' and R may be the same or ditferent substituents andare selected from the group consisting of hydrogen, methyl, ethyl,propyl and phenyl; X is an amino radical or amino acid; and acid saltderivatives of the same, such as acetic, hydrochloric, sulfuric, etc.;and the method of preparation thereof, The intermediate (i)-9-[ri-(3a-amino 2a hydroxy/)cyclopentyl] 6 dimethylaminopurine, or any ofits corresponding 6-substitnted derivatives, leads to the preparation oftwo diasterioisomers when condensed with the appropriately blocked L-amino acid. The separation of these diasterioisomers results in theisolation of antimicrobial and antitumor agents.

The invention described herein was made in the course of work under agrant or award from the Department of Health, Education, and Welfare.

This invention relates to a new class of chemical compounds,(i-)9-[-(3a-amino-Za-hydroxy)cyclopentyl]-6- substituted-purines havingthe general formula:

wherein R is a substituent selected from the group consisting ofhydroxyl, mercapto, methyl mercapto, halogen (chlorine, bromine, uorineand iodine) and substituted amino:

wherein R and R" may be the same or diiferent substituents and areselected from the group consisting of hydrogen, methyl, ethyl, propyland phenyl; X is an amino radical or amino acid; and acid saltderivatives of the same, such as acetic, hydrochloric, sulfuric, etc.Although the invention is described in detail with reference to thedimethyl substituted aminopurine and selected derivatives thereof, itwill be readily understood that the same general ow scheme may befollowed to produce compounds with ditferent radicals substituted in the-position and different derivatives thereof.

DESCRIPTION OF THE PRIOR ART Puromycin, an antibiotic with antitumoractivity [B. L. Hutchings, Chem. Biol. Purz'nes, CIBA Found. Symp., 1956777 (1957)] has been found to inhibit protein biosynthesis in a widevariety of organisms. However, since puromycin is a ribonucleosideantibiotic, it is easily cleaved hydrolytically or enzymatically in vivothus lessening its effect as a chemotherapeutic agent. [P. M. Rull, H.Weinfeld, E. Carroll, and G. B. Brown, J. Biol. Chem., 220 439 (1956)and B. R. Baker, Design at Active-Site- Directed Irreversible EnzymeInhibitors. The Organic Chemistry of the Active Site, John Wiley andSons, Inc., New York, N.Y., 1967, pp. 79, 93.] Toxic manifestations,including renal lesions, have precluded the use of puromycin in thetreatment of human or animal infectious diseases or neoplasms. [D.Nathans in Antibiotics I, Mechanism of Action, D. Gottlieb and P.D.Shaw, Eds., Springer-Verlag, New York, N.Y., 1967, pp. 259-277.] Thenephrotic syndrome results from the formation of toxic puromycinmetabolites formed after administration of the antibiotic.

SUMMARY OF THE INVENTION cancer, etc.) die from a lack of protein. As aresult, the.

drug can be used to treat infectious diseases. The compounds represent anew concept in the design of puromycin analogs. The molecule retains allof the moieties necessary for growth inhibition, and at the same time itlacks the moieties which are responsible for toxicity. The greatestadvantage lies in the use of these drugs against non-bacterialinfections such as protozoan and trypanosome diseases, since mostantibiotics are not active against non-bacterial organisms.

DETAILED DESCRIPTION OF THE INVENTION FIGS. 1 and 1A together representa ow sheet leading to (1) -9- 3 a-amino-Za-hydroxy cyclopentyl]-6-substituted-aminopurines and derivatives thereof.

The carbocyclic analog of puromycin g was synthesized by the route shownin FIGS. 1 and 1A (Structures -l depict only one enantiomer of theracemic form actually obtained.) Cyclopent-Z-enone ethyleneketal wasreacted with N-bromosuccinimide in ether-water by a modification of theprocedure of Guss and Rosenthal in which NaHco3 was used to buffer thereaction mixture. Cyclopent-2- enone ethyleneketal (37.85 g., 0.300mole), N-bromosuccinimide (53.40 g., 0.300 mole), NaHCO3 (4.20 g., 50.0mmoles), Et2O (240 ml.), and H2O (240 ml.) were stirred vigorously for6.5 hours, at which time all of the solid had disappeared and the pH wasapproximately 7. The aqueous layer was saturated with NaCl, and the Et2Olayer then separated. The aqueous layer was extracted with additionalEt2O (2X 100 mL). The combined EtZO layers were washed with saturatedNaCl and dried (CaSO4). When bromohydrin 4 was immediately treated withNaOH in reuxing benzene, epoxide 6- oxabicyclo[3.1.1]hexan-2-oneethyleneketal, was formed in 73% yield (from as a stable colorlessliquid. The epoxide was opened with sodium azide in dioxane by themethod of Vander Werf et al., and the resulting azide reducedcatalytically to crystalline trans-3-amino-2-hydroxycyclopentanoneethyleneketal Z in 76% yield (from A solution of (2.84 g., 20.0 mmoles)in dioxane (40 ml.) was brought to reflux. A solution of sodium azide(1.63 g., 25.0 mmoles) in H2O (10ml.) was added to the refluxingsolution over 1.0 hour. The resulting mixture was reuxed with vigorousstirring for 48 hours. The dioxane layer was separated from the aqueouslayer, and the aqueous layer was extracted with additional dioxane (50ml.) The combined dioxane layers were washed with saturated NaCl (50ml.) and dried (CaSO4). This material was dissolved in absolute EtOH (75ml.) and shaken with platinum oxide (100 mg.) under hydrogen (50 p.s.i.)for 20 hours. After ltration, the EtOH was evaporated, leaving paleyellow solid (2.91 g.) which IR showed to contain no azide.Crystallization from PhH gave 7 as pale yellow crystals (2.41 g.), m.p.97-98.

The purine moiety was formed via a standard method. Condensation of Zwith -amino-4,6-dichloropyrirnidine, followed by ring-closure of theresulting crude pyrimidine with triethylorthoformate in the presence ofethane sulfonic acid gave the 6-chloropurine Q, trans-3-(6-Chloro-9-purinyl) 2 hydroxycyclopentanone Ethyleneketal, in `84% yield (from 1)as follows: A solution of Z (14.04 g., 88.20 mmoles),5-amino-4,6-dichloropyrimidine (14.47 g., 88.20 mmoles), andtriethylamine (37 ml., 265 mmoles) in n-BuOH (160 ml.) was reuxed undernitrogen for 44 hours. Evaporation (50, 0.2 mm.) left a brown oil (42g.) containing which was shaken with triethylorthoformate (200 rnl.) fora few minutes. The white solid Et3NH+C1* (10.82 g., 89%) did notdissolve and was removed by filtration. Ethane sulfonic acid (1.0 g.)was added to the filtrate, and it was stirred at ambient temperature forhours. At this time solid had formed. Hexane (200 ml.) was added, andthe mixture was Cooled. The tan solid was collected and washed withhexane (100 ml.), giving crude l (24.42 g.), Crystallization from EtOAc(500 ml.) gave tan needles (22.03 g.), m.p. 165- 167.

The 6dimethylaminopurine 1Q,trans-3-(6-Dimethylamino-9-purinyl)-Z-hydroxycyclopentanoneEthyleneketal, was formed in 75% yield when Q was treated with refluxingaqueous dimethylamine. A solution of Q (3.85 g., 13.0 mmoles) in aqueousdimethylamine (100 ml.) was refluxed 3.0 hours. The solution wasconcentrated to 20 ml. and extracted with EtOAc (3 100 mL). The EtOAcextract was dried (CaSO4) and concentrated to 50 ml. White crystals of1Q were collected (2.99 g.), m.p. 156- 158.

Other substitutions may be made in the -6- position according to theprocedures described in the papers of H. I. Schaeffer and R. Vince,Journal of Medicinal Chemmy, 8, 33, (1965) and H. J. Schaeffer, D. vogeland R.'

Vince, Journal of Medicinal Chemistry, 8, 502, (1965). Typicalalternative substituents include hydroxyl, mercapto, methyl mercapto,chlorine, bromine, uorine, iodine and amino groups other than thedi-methyl-amino radical illustrated. The substituents of thecli-substituted amino group may be the same or different and includehydrogen, methyl, ethyl and phenyl radicals.

Oxirne Q, trans-3-(6-Dimethylamino-9-purinyl)2-hydroxycyclopentanoneOxime, was synthesized directly from Q under what appear to be uniqueconditions for oxime-formation. To a stirred mixture of (10.00 g., 32.8mmoles), hydroxylamine hydrochloride (11.38 g., 164 mequiv.), and H2O(140 ml.) was added 2 N HCl (approximately 28 ml.) dropwise until the pHwas 1.0 (measured on a meter), and all solid had dissolved. The solutionwas stirred at 70-75 for 3.0 hours. The pH of the hot solution was thenadjusted to 6.5 `with 6 N NaOH, and white solid began to precipitate.The mixture was cooled (5) for several hours. The solid was collected,washed With H2O (50 ml.), and air-dried, giving (8.95 g., 99%), m.p.202-204 dec; tlc indicates a mixture of syn and anti oximes. Sinceattempted separation by chromatography or recrystallization resulted inconsiderable losses, the mixture was acetylated without separation, asolution of 1 1 (8.95 g., 32.4 mmoles) in Ac2O (200 ml.) was stirred at6065 for 4.25 hours. Evaporation left a tan solid which was crystallizedfrom EtOAc, giving a chromatographically homogeneous diacetyl derivativeE as white crystals (8.16 g., 70%), m.p. 152-154.

The O-acetyloxime 1 2,trans-2-Acetoxy-3-(-dimethylamino-9-purinyl)cyclopentanoneO-Acetyloxime, was reduced to a mixture of aminoalcohols with diboranein tetrahydrofuran (THF) by a modification of the procedure yused byFeuer and Braunstein for the conversion of cyclohexanone O-acetyloximeto cyclohexylamine. To a stirred, cooled (0-5) solution of 1 2 (4.37 g.,12.1 mmoles) in dry THF (100 ml.) was added a 1M solution of BH3 in THF(44.0 ml., approximately 130 mequiv. hydride) over a period of 1.0 hourunder a nitrogen atmosphere. The solution was stirred for an additional3.0 hours at 045, and then for 12.0 hours at ambient temperature. Aftercooling the reaction solution (ice bath), H2O (8.3 ml.) was added. TheTHF was evaporated (25, 0.5 mm), and the residue stirred with 2N HC1(110 ml.) at ambient temperature for 3.5 hours. After evaporation, theresidue was dissolved in MeOH (50 ml.) and passed slowly through acolumn of 100 ml. of Amberlite IRA-400 resin (OH*) packed in MeOH. Thebasic eluent (500 ml.) was evaporated, leaving a yellow, gummy solid(3.26 g.), which TLC indicated to be largely a mixture of aminoalcoholsand numerous minor contaminants. The amino-alcohols could not heseparated by preparative tlc and appeared to carbonate on standing. Thisgummy solid was dissolved in Ac2O and stirred at 60 for 1.75 hours.Evaporation left a brown glass (3.74 g.) which was chromatographed on acolumn of silica gel (200 g.) packed in CHC13. Elution with 2%MeOH-CHC13 (2 l.) gave a pale yellow glass (290 mg), which crystallizedfrom EtOAc-hexane to a white solid (64 mg), m.p. 181-183. Elution with3% MeOH-CHC13 (3 1.) gave l? ,(l)9[(3a-Acet amido 2ahydroxy)cyclopentyl] 6 dimethylaminopurine, as a pale yellow glass (2.09g.), which crystallized from EtOAc (1.65 g., 45% from m.p. 151-152".

Continued elution with 3%- MeOH-CHCla (1.51) gave fractions containingboth l and (124 mg., 3%), followed by fractions containing only(1)-9-[a-(3a- Acetamido-Z-hydroxy)cyclopentyl] 6 dimethylaminopurine, asan amorphous white solid (247 mg). Crystallization of this solid fromEtOAc gave 1i as white crystals (159 mg., 4%), m.p. 171-171.5.

The diacetyl derivatives 13 a and 1 4@ were obtained when the mixturefrom the diborane reduction of 1.00 g. (2.78 mmoles) of 1 2 was stirredin Ac20 (5` ml.)pyridine (10 ml.) at ambient temperature for 18 hours.

Evaporation left a brown glass (800 mg.) which was chromatographed on acolumn of silica gel (50 g.) packed in CHC13. Elution with 1% MeOH-CHC13(700 ml.) gave 14 a as a colorless glass (154 mg.). Crystallization fromEtOAchexane gave white solid (32 mg., 3%), m.p. 207.5-2l0 dec.

Elution with 2% MeOH-CHC13 (850 m1.) gave a yellow glass whichcrystallized from EtOAc giving 1 3 a as tan crystals (377 mg., 39%),m.p. 194.5-196".

The N-acetyl derivative 1 3 (730 mg., 2.40 mmoles) was reiiuxed in asaturated aqueous solution of Ba(OH)2 (approximately 0.5N, 40 ml.) for3.0 hours. The solution was dil-uted with EtOH (50 rnl.) and treatedwtih excess Dry Ice. The BaCO3 was removed by filtration through Celite.Evaporation left a white solid (740 mg.), m.p. ISO-182 dec., which wasresolidied from EtOH-EtzO, giving E, Acetic Acid Salt of (i)9-[-(3-Amino-2ahydroxy)cyclopentyl]6dimethyl aminopurine, as a whitepowder (685 mg., 89%), m.p. 184-186 dec.

The procedure of Baker and Joseph was used to prepare the 2,3carbamateof the puromycin aminonucleoside 1 8 (i )9 3a-Amino-2a-hydroxycyclopentyl] 6dimethylaminopurine 2', 3Carbamate. A solution of (100mg., 0.310 mmole) and triethylamine (0.13 ml., 0.93 mmole) in dimethylformamide (DMF) (5 ml.) was cooled to 5. Carbobcnzoxy chloride (0.05ml., approximately 0.5 mmole) was added, and the solution stirred atambient temperature for 1.0 hour. H2O (20 ml.) was added and the oilwhich formed was extracted into CHC13 (3X 20 ml.). The CHC13 extractswere dried (CaSO4) and evaporated, leaving a colorless glass (96 mg),which TLC in solvent B showed to be a mixture of 11 and 18 inapproximately equal amounts. When the mixture of 11( and 1 8 was treatedwith NaOMe in DMF as described below, was isolated in 74% yield (from1Q).

The carbocyclic aminonucleoside analog 1 6 was coupled toN-benzyloxycarbonyl-p-methoxyphenyl L alanine by two methods: (A) thedicyclohexylcarbodiimide-N-hydroxysuccinmide method and (B) amodification of the mixed anhydride method suggested by Anderson et al.The resulting carbobenzoxy blocked diastereomers 23, 9{R[SR-(Benzyloxycarbonyl p methoxyphenyl Lalanylamino)-2Rhydroxy]cyclopentyl} 6 dimethylaminopurine, and 19 a,9-{S-[3S-(Benzyloxycarbonyl p methoxyphenyl-L-alanylamino) 2Shydroxy]cyclopentyl}6di methylaminopurine, (97% by method A, 77% bymethod B) could not be separated. Alternatively, other amino acidderivatives of 1 6 may be prepared by analogous procedures.

Method A A solution of 1g (145 mg., 0.450 mmole) in MeOH was passedslowly through a column of 10 ml. of Amberlite IRA-400 resin (OH) packedin MeOH. Evaporation left the free amine as a white solid (118 mg.,0.450 mmole), m.p. 153-154, which was dissolved immediately in DMF (5mL), along with N-benzoyloxycarbonyl-pmethoxyphenyl-L-alanine (155 mg.,`0.472 mmole) and N-hydroxysuccinimide (54.3 mg., 0.472 mmole). Thesolution was cooled to 0, and DCC (97.4 mg., 0.472 mmole) was added.After stirring at 0 for 30 minutes, the solution was allowed to stir atambient temperature for 20 hours. The mixture was filtered, thedicyclohexylurea washed with EtOAc (20 m1.), and the combined filtrateevaporated. A solution of the residue in EtOAc (5 ml.) was cooled at 0and then filtered. The liltrate was diluted to ml. with EtOAc, extractedwith H2O (2.5 mL), then one-half saturated NaHCO3 (2.5 mL), then H2O (2X5 ml.). Evaporation of the dried EtOAc solution left a mixture of ga and19 a as a white solid foam (250 mg., 0.436 mmole).

Method B A solution of triethylamine (61.3 mg., 0.606 mmole) in dry THF(10 ml.) was cooled to -10. Ethyl chlorocarbonate (72.4 mg., 0.667mmole) was added. The solution was stirred for 1 minute, and thenN-benzyloxycarbonyl-p-methoxyphenyl-L-alanine (200 mg., 0.606 mmole) wasadded andthe mixture stirred at -10 for 10 minutes. A sample of the freeamine obtained by resin treatment of 1E as in method A (159 mg., 0.606mmole) was dissolved in DMF, the solution cooled to y 10 and added tothe mixed anhydride. The resulting mixture was stirred at -l0 for 1.0hour and then stored at 4 for an additional 24 hours. The mixture wasiiltered, and the solid collected washed with additional DMF (5 ml.).Evaporation of the combined tiltrates left a colorless glass which wastriturated with H2O (5 ml.), evaporated to dryness, and chromatographedto prep TLC on a plate developed in solvent C. Extraction of the majorband with 20% MeOH-CHC13 (other bands were unreacted starting materials)gave a mixture of 2 a and lia as a solid foam (266 mg., 77%), Rf, IR,and NMR identical with the mixture obtained from method A.

The crude mixture of aminoalcohols resulting from the diborane reductionof 3.00 g. (8.32 mmoles) of 1g was reacted directly with the mixedanhydride of N-benzoyloxycarbonyl-p-methoxyphenyl-L-alanine in thismanner, giving a mixture of ga and after chromatography, in 30% yield(from 1 2); Rf, IR, NMR identical with the samples described above. Themixture of aminoalcohols from the reduction of Q was also coupled toN-benzoyloxycarbonyl-p-methoxyphenyl-L alanine by method A, giving ayield of ga and Qa, after chromatography, comparable with the overallyield via 18 and 1 6. Following hydrogenolysis of the carbobenzoxygroup, separation of diastereomers g, 6Dimethylamino-9-{R-[2R-hydroxy-3R- (pmethoxyphenyl-L-alanylamino)]cyclopentyl}purine, and E, 6Dimethylamino-9-{S-[2S-hydroxy3S-( p methoxyphenyl-L-alanylamino)]cyclopentyl}purine by chromatography was possible. A mixture of 2 a and1 9 a prepared by method A V(250 mg., 0.436 mmole) was dissolved inglacial AcOH (15 ml.) and shaken with 10% Pd-C 125 mg.) under hydrogen(1 atm.) for 10 minutes, by which time hydrogen uptake had ceased. Themixture was filtered through Celite and the Celite was washed withadditional AcOH 10 ml.). Evaporation of the combined filtrate and wash(35, 0.5 mm.) left a colorless glass. A solution of this glass in MeOHwas passed slowly through a column of 10 ml. of Amberlite IRA400 resin(OH) packed in MeOH. The basic MeOH eluent (250 Inl.) was evaporated,leaving a mixture of 2 and 1 9 as a white solid (176 mg., 92% m.p.194-196.

The diastereomers g and Q were separated by prep TLC (50-70 mg. perplate) in solvent D. The two bands were each stirred for 18 hours with20% MeOH-CHC13, filtered, and evaporated, giving almost quant recoveryof the pure diastereomers as colorless glasses.

Hydrogenolysis of a mixture of 2 a and 19 a prepared by method B gave a91% yield of 2 and Q. The two coupling methods resulted in samples of gand with identical optical purities. The mass spectra of 2 and E arealmost identical, differing slightly only in the relative intensities ofsome ions.

RESULTS AND DISCUSSION Antimicrobial testing of diasteriomers g and 1 2revealed that one isomer was completely inactive while the otherexhibited growth inhibition on the same order of magnitude as puromycin.The absolute stereochemistry of the active isomer has tentatively beenassigned that of struc ture 2 on the basis of its biological activityand in accordance with the stereochemistry of puromycin. The minimuminhibitory concentrations by a twofold serial dilution test in broth forpuromycin and the carbocyclic analog 2, respectively, are as follows(mM.): Staphylococcus aureus (NRRL B-3l3), 0.244 and 0.244; Bacillussubtilis (NRRL B-545), 0.030 and 0.060; Klebsiella pneumoniae (NRRL B-l17), 0.485 and 0.485; Escherichia coli (NRRL B-210) 0.060 and 0.120. Agrowth curve for S. aureus in the presence of ditferent concentrationsof puromycin or the carbocyclic compound shows a lag period with bothcompounds when the concentrations are lower than those required forcomplete inhibition. ,Such a lag period is consistent with the mechanismof action of puromycin since the antibiotic would be expected to beconsumed as it is incorporated into the growing peptide chains.

The aminonucleoside E was tested for nephrotoxicty in rats at a dose of33 Ing/kg. under the same conditions that are .required for puromycinaminonucleoside to cause severe nephrotic syndrome at a dose of mg. Nonephrotoxicity was observed even after 17 days of treatment with 1 6.

The novel puromycin analog g provides a molecule with the structuralfeatures required for puromycin-like antimicrobial activity. Thus, theribofuranosyl ring can be replaced with the more hydrolytically stablecyclopentane ring without loss of activity. In addition, the removal ofthe hydroxymethyl moiety is not detrimental to activity and at the sametime provides g with a resistance to kinase activity upon release of theaminonucleoside. This resistance to phosphorylation circumvents thenephrotic syndrome associated with puromycin aminonucleoside. Thiscarbocyclic analog and others which are under preparation are beingevaluated for in vitro inhibition of protein biosynthesis in an attemptto explore the requirements for binding to ribosomes. Preliminarystudies with g and 1 9 on an E. coli ribosomal system are consistentwith the antimicrobial activities.

It is apparent that many modilications and variations of this inventionas hereinbefore set forth may be made without departing from the spiritand scope thereof. The specific embodiments described are given by wayof example only and the invention is limited only by the terms of theappended claims.

The embodiments of the invention in which an exclusive property orprivilege is claimed are defined as follows:

1. (i)9[(3a-Amino 2a hydroxy)cyclopentyl]6 substituted-purine having thegeneral formula:

wherein R is a substituent selected from the group consisting ofhydroxyl, mercapto, methyl mercapto, chlorine, bromine, fluorine andiodine and substituted amino:

References Cited UNITED STATES PATENTS 3,016,378 1/1962 Roch 260-252DONALD G. DAUS, Primary Examiner A. M. T. TIGHE, Assistant Examiner U.S.Cl. X.R.

gyg UNITED STATES PATENT OFFICE CERTIFICATE 0F CORRECTION patent Nc.3,25,'541 Dated July 25. 1974 Inventor Robert Vince It is certifiedvthan; error appears in the above-identifi.=.d patent and that saidLetters Patent are hereby corrected as shown below:

l'- l l "l Inn the heading-of the drawings, the title should be: (i) -9[f (Bcf-AMINO-ZHYDROXY)CYCLOPENTYL] --SUBSTITUTED- PURINES ANDDERIVATIVES THEREOF Column 2, line 16, 6- should be 6.

Column 3, line S75, 8 should be underscored.

`Column 4, line l, 8 should be underscored.

I Column 5 line 42 9{R should be 9- fR I Column 5, line 43, L Should beitalicized.

Column l 7 liney l2 l5 mg should be 15ng/kg.

Signed and sealed this 5th day of November 197A.

(SEAL) Attest:

MCCOY M, GIBSON JR. C. MARSHALL DANN .Ai-,testing Officer Commissionerof Patents

